Preliminary studies of cell culture strategies for bioprocess development based on HEK293 cells

Liste‐Calleja, Leticia; López‐Repullo, Jonatan; Lecina, Martí; Cairó, Jordi J.

doi:10.1186/1753-6561-7-s6-p52

Cited by 1 publication

(1 citation statement)

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“…The initial experiments in 2 L stirred-tank bioreactors operated in batch mode demonstrate the impact of increased culturing times, medium saturation, and higher cell densities on AAV production. These profiles are comparable to other HEK 293 cultures operated batchwise (Koh et al, 2009;Gálvez et al, 2012;Liste-Calleja et al, 2013). The potential limitation of nutrients and/or the accumulation of toxic by-products had a negative effect on specific productivities, this being more pronounced at higher VCC TOT with a 5-fold lower specific production in BR5 (VCC TOT = 5.57 × 10 6 cells/mL) when compared to BR2 (VCC TOT = 2.61 × 10 6 cells/mL).…”

Section: Discussionsupporting

confidence: 78%

AAV process intensification by perfusion bioreaction and integrated clarification

Mendes

Fernandes

Pineda

et al. 2022

Front. Bioeng. Biotechnol.

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Adeno-associated viruses (AAVs) demand for clinical trials and approved therapeutic applications is increasing due to this vector’s overall success and potential. The high doses associated with administration strategies challenges bioprocess engineers to develop more efficient technologies and innovative strategies capable of increasing volumetric productivity. In this study, alternating tangential flow (ATF) and Tangential Flow Depth filtration (TFDF) techniques were compared as to their potential for 1) implementing a high-cell-density perfusion process to produce AAV8 using mammalian HEK293 cells and transient transfection, and 2) integrating AAV harvest and clarification units into a single step. On the first topic, the results obtained demonstrate that AAV expression improves with a medium exchange strategy. This was evidenced firstly in the small-scale perfusion-mocking study and later verified in the 2 L bioreactor operated in perfusion mode. Fine-tuning the shear rate in ATF and TFDF proved instrumental in maintaining high cell viabilities and, most importantly, enhancing AAV-specific titers (7.6 × 104 VG/cell), i.e., up to 4-fold compared to non-optimized perfusion cultures and 2-fold compared with batch operation mode. Regarding the second objective, TFDF enabled the highest recovery yields during perfusion-based continuous harvest of extracellular virus and lysate clarification. This study demonstrates that ATF and TFDF techniques have the potential to support the production and continuous harvest of AAV, and enable an integrated clarification procedure, contributing to the simplification of operations and improving manufacturing efficiency.

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Section: Discussionsupporting

confidence: 78%